We propose to integrate two new technologies to develop an radical new approach to making, utilizing and reading DNA arrays. The first is construction of an array of single molecule probes arranged at nanometer- scale spacing using self-assembly. The approach is based on the DNA nanoassemblies, but in this proposal we will develop methods for placing unique, addressable tiles at each point in an array. The second is a nanoscale readout using molecular recognition imaging based on antibody-modified atomic force microscope probes with a real-time detection scheme for anti-body binding during topographic imaging. This technology enables robust scanning of the chemical composition of nm-scale structures, and is capable of reading out 30,000 sites per minute with current technology. Taken together, these two technologies should enable (a) Low cost fabrication: A nM-scale synthesis yields ca. 100 trillion arrays; (b) Intrinsic single molecule detection with a target density limited only by the binding kinetics; (c) Remarkable potential probe densities - a billion sequences could be fitted onto an array of centimeter dimensions; (d) Solution phase processing - current arrays of ca. 10 micron size (potentially containing 250,000 probes) are soluble. The arrays are only deposited onto a surface for the final readout step, (e) The potential for picogram (or much smaller) amounts of sample, because the volume occupied by the arrays is very small indeed. Our goals here are to test the concept, quantify the performance and lay out a path for production of practical devices. We will also develop and test labeling schemes, both for efficient readout, and for SNP detection. Our test array will incorporate SNPs from strains of Yersina pestes. If successful, a subsequent full proposal will address extension of the technology to very large arrays and to very small sample volumes. We hope that this combination of addressable nm-scale "molecular pegboards" combined with single molecule readout will eventually prove useful in many areas of analysis and as a basis for assembling molecular complexes for synthetic biology. [unreadable] [unreadable]